Project Summary/Abstract Epithelial-mesenchymal transitions (EMTs) are highly regulated dynamic processes in which cells in stable epithelia acquire the ability to migrate and organize new tissues. EMTs are essential for establishment of tissues layers in developing embryos and for the development of many different organs, and disrupted EMTs can cause birth defects. In the adult, abnormal EMTs can cause fibrosis in the kidney, liver and lung, and they can also drive tumor progression and metastasis. Despite their importance, the dynamics and regulation of the cellular events of mammalian EMTs in vivo are poorly understood. The mouse gastrulation EMT provides an unparalleled context to combine the tools of genetics, imaging and cell biology to define the cellular, tissue and mechanical mechanisms that control cell behavior during mammalian epithelial-to- mesenchymal transitions in vivo. The mouse gastrulation EMT, which generates the three body layers of the animal, provides a uniquely advantageous context to dissect the molecular and cellular processes that regulate an EMT. The gastrulation EMT is relatively rapid: individual cells move from the epithelium to the mesenchymal layer in less than an hour. Signaling pathways and transcription factors that are required for this EMT have been identified, and the EMT can be visualized using fluorescent transgenic reporters in real time. This proposal uses a novel set of mouse mutations to define the cell biological events required for the EMT. Genetic and cell biological experiments will test the novel hypothesis that a self-organizing network of interactions among a set of apical epithelial proteins controls the stochastic ingression of cells during gastrulation. Experiments will test whether FGF signaling directly promotes the cell biological changes that drive cells to exit the epithelium, in addition to its established role in the regulation of gene expression. The final step of the EMT is the organized and directed migration of the newly formed mesenchymal cells to generate the organs of the animal. Experiments will test the hypothesis that regulated migration of mesoderm cells drives elongation of the anterior-posterior body axis and that directed mesoderm migration depends on the Striatin Interacting Phosphatases and Kinases (STRIPAK) protein complex. This work will provide the first dynamic analysis of a genetic network that controls the cellular events of a mammalian EMT in vivo and will provide a foundation for understanding other normal EMTs, as well as how aberrant EMTs cause human disease.